The tRNAs for Asp, Asn, His and Tyr contain a hypermodified purine known as queuine in the first position of the anticodon. In order to determine if queuine was synthesized by higher mammals, germfree mice were fed a chemically defined diet that lacked queuine for nine tRNA half lives and the liver tRNA was then examined for queuine content and compared with a control group of germfree mice fed sterilized commercial mouse chow. The queuine content of tRNAAsn and tRNAHis decreased five-fold in the chemically defined diet mice but there was no decrease in tRNAAsp and tRNATyr. These experiments suggest that there is a dietary requirement for queuine in rodents and probably other mammals. These experiments also indicate that there is a mechanism for salvaging and recycling queuine but the study has given no clues to the physiological role of queuine. The objectives of this proposal are to determine the effects of long-term deprivation of queuine on tRNA and on health. Studies entailing deprivation of queuine will be carried out to see if it is possible to totally eliminate queuine from the tRNA or if some mechanism that obviates the requirement for dietary queuine comes into play during queuine deprivation. Weaned mice grow and show no adverse effects on a queuine deficient diet, but there is a marked decrease in reproduction rate, and mice born into a queuine deficient environment have decreased lifespans and experience a bizarre sudden-death syndrome. The chemically defined diet will be supplemented with queuine to see if these two dyscrasias are prevented. We also intend to study the enzymatic mechanism by which the body salvages and recycles queuine. Queuine appears to play a role in normal reproduction (fetal serum and amniotic fluid contain large amounts of it). The queuine content of tRNAs of malignant cells is less than normal. Queuine-containing tRNA has been shown to control the synthesis of at least one enzyme, asparagine synthetase. The purpose of this proposal is to study the effect of dietary queuine deprivation on these phenomena and to fill the gaps in our basic understanding of queuine metabolism.